Reduced Representation Bisulfite Sequencing provides a powerful method to efficiently analyze DNA methylation at the single nucleotide level without the higher costs associated with whole genome bisulfite sequencing. By cutting the genome using the restriction MspI enzyme (CCGG target sites) followed by size selection, DNA is enriched to represent CpG-rich regions (including CpG islands), in which DNA methylation marks are typically found. Thus, RRBS provides a cost-effective method for analyzing DNA methylation by reducing the part of the genome that actually needs to be sequenced and focusing on relevant CpG islands.

They love it! The new Diagenode Premium RRBS Kit makes it easy to use RRBS cost-effectively and with high throughput, using early sample pooling and multiplex sequencing. Most importantly, the method provides an improved coverage of up to 4 million CpGs for the human genome. We successfully used this protocol on more than 1,000 samples comprising of six different species, various cancers, FFPE and lowinput samples. Paul Datlinger and Christoph Bock, CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria

Accurate determination of DNA methylation level

Excellent results were obtained using Diagenode’s Premium RRBS kit: almost 90% alignment rate, 4.1 million CpGs covered and bisulfite conversion rates around 99.5% for all samples. DNA methylation percentages in the region of IGF2 obtained with the Diagenode’s Premium RRBS Kit. Two human cell lines were analyzed: Gm12878 and MCF7. The MCF7 cell line was studied in duplicates. Each peak represents the DNA methylation percentage at one CpG. The methylated CpGs are shown in red and the unmethylated CpGs in grey.

How it works

By cutting the genome using the restriction enzyme MspI (CCGG target sites) followed by size selection, DNA is enriched to represent CpG-rich genomic regions (including CpG islands, CpG island shores, enhancers, and other gene-regulatory elements), which are particularly relevant for epigenetic regulation. Similar to exome-sequencing for mutation discovery, the RRBS protocol enriches for some of the most interesting target regions and thereby achieves a reduction in sequencing cost of a factor of 10-20 compared to whole genome bisulfite sequencing.

The new Diagenode Premium RRBS Kit makes it easy to use RRBS cost-effectively and with high throughput, using early sample pooling and multiplex sequencing. Most importantly, the method provides an improved coverage of up to 4 million CpGs for the human genome. We successfully used this protocol on more than 1,000 samples comprising of six different species, various cancers, FFPE and lowinput samples.

Paul Datlinger and Christoph Bock, CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria

Our laboratory has used RRBS sevice of Diagenode on murine and human samples. The service was impeccable in each phase, from the sample preparation to bionformatic analysis because it was always customer-oriented. I highly recommend my colleagues to use the RRBS service from Diagenode.

Using the Premium RRBS kit we quickly established the RRBS library preparation in our group. Diagenode's support on wet-lab matters as well as on bioinformatics was exceptionally customer-oriented and close, accelerating the establishment of the method.

Our lab has used Diagenode’s Premium RRBS kit on rat brain samples. The protocol is understandable, logical, well-written and is easy to follow. It is a complicated, but ingenious kit. I found it fantastic that I could ask questions from the company, and their answers were really useful. We were able to construct a library, which we ran on BioAnalyzer, and the results looked very nice and ready to be sequenced. I would definitely recommend my colleagues to use the Premium RRBS kit from Diagenode.

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